It remains unclear how are TCR and its co-receptor signals controlling lymphocyte development and differentiation regulated intracellularly. We previously demonstrated that adaptor molecule Cbl is involved in organizing TCR signals in thymocytes, and its deficiency leads to an enhanced CD4+ thymocyte development in vivo. Using gene knock-out (ko) mice, we now demonstrate that Cbl-b, a member of Cbl family proteins, is required for the establishment of CD28 dependence of T cell activation, and that in the absence of Cbl-b mice become highly susceptible to the induction of autoimmune diseases. Further biochemistry analysis indicates that the Vav signaling pathway is significantly enhanced in the mutant T cells, suggesting that Cbl-b regulates the CD28 dependence through inhibiting Vav activation by TCR signals. Our results for the first time show that an adaptor molecule is involved in the coordination of TCR and its co-receptor signaling, and suggest further that dysregulation of Cbl-b and Vav signaling pathways might contribute to the development of autoimmune diseases in humans.